We have established a stable human cell line, termed HPL-A3, by co-transfection of a human pregnane X receptor (hPXR) expression vector and a reporter plasmid (p3A4-hPXRE-Luc) containing a luciferase gene and a promoter/enhancer region of the human cytochrome P450 3A4 (CYP3A4) gene into a human hepatomaderived cell line, HepG2. We then examined the usefulness of HPL-A3 for a chemically activated luciferase expression (CALUX) assay of human CYP3A inducers. The induction of CALUX in HPL-A3 by

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... activators, including rifampicin, occurred in time-and concentration-dependent fashions, whereas no such induction was observed using rat/mouse PXR activators, such as pregnenolone-16α-carbonitrile and dexamethasone. The hPXR activator-mediated induction of CYP3As, especially CYP3A4, was observed at levels of both mRNA and enzyme activity. Furthermore, there were positive correlations between chemical-mediated inductions of CALUX and CYP3A4 mRNA levels. In addition, the induction of CALUX by dihydropyridine calcium channel blockers, which are known to act as CYP3A inducers in rats, was observed in HPL-A3 cells. Interestingly, expression levels of not only hPXR but also of vitamin D receptor (VDR), a transcription factor that positively regulates CYP3A subfamily genes, were significantly increased in HPL-A3 cells compared with those in the parental cell line, HepG2. Consequently, VDR ligand (1,25-dihydroxyvitamin D 3 )-mediated inductions of CALUX and CYP3A4 mRNA were observed in the cells. These findings verified the usefulness of HPL-A3 for the screening of CYP3A inducers, which can activate the hPXR and/or hVDR.